Abstract The complement system is not only an integral arm of innate immunity but also significantly contributes to the menacing severity of infection-associated inflammation. The life-threatening inflammatory syndrome of acute respiratory distress syndrome (ARDS) frequently arises from uncontrolled bacterial pneumonia imposing persistent public health burdens. These disorders involve activation of the complement system with excessive generation of the complement anaphylatoxin, C5a, and therapeutic disruption of these processes could provide protection from tissue-destructive inflammation. C5a ligates with its homologous receptors, C5aR1 and C5aR2, encoded by two adjacent genes. The functional roles of C5aR1 and C5aR2 across different tissues are not entirely clear, and the data for C5aR2 is controversial. In our preliminary work, we have generated a mouse strain with a ~12.6 kb deletion of both C5aRs (C5aR1/2-/- DKO) by CRISPR/Cas9 gene editing, another mouse strain for reporting and conditional deletion of C5aR2 (C5aR2LacZ; C5aR2flox) and we have access to conditional C5aR1 mice (C5aR1flox). These novel tools will facilitate the testing of our central hypothesis that a functional cooperation of C5aR1 on neutrophils with C5aR2 on type II alveolar epithelial cells determines the development and progression of lung injury and pneumococcal pneumonia. We will focus on three specific aims: (1) Based on our preliminary findings with bone marrow chimeric mice, the expression and functional relevance of C5aR2 in type II alveolar epithelial cells (AECIIs) will be studied including endpoints such as alveolar-capillary barrier disruption, signatures of cytokines/chemokines and epithelium-leukocyte cross-talk during lung injury and pneumococcal pneumonia. In addition, the presence of C5aR2 will be compared to C5aR1 in normal human lung tissues. (2) We will characterize the roles of C5aRs in Ly6G+ neutrophils during lung injury and pneumococcal pneumonia using mice with neutrophil-specific deletion of C5aR1 and C5aR2. To elucidate potential effects of C5aR1 on subsets of lung invading neutrophils and their maturation states, we will pursue single-cell RNA-Seq studies using a novel droplet-based high-throughput method. (3) We will investigate the functional synergisms, redundancies and diversities of C5aRs by comparing the inflammation phenotypes of C5aR1/2-/- DKO to C57BL/6 (Wt), C5aR1-/- and C5aR2-/- mice during lung injury and pneumococcal pneumonia. Neutrophils and AECIIs isolated from the aforementioned pneumonia experiments will be used for comparing the whole transcriptomes of these four mouse strains in deep sequencing studies (bulk RNA-Seq) focusing on elucidation the molecular pathways of neutrophil-AECIIs interactions and a search for novel C5aR1/2-regulated genes. These investigations will aim to advance the current concepts and resolve ongoing controversies regarding the functional antagonism or operational synergism of the C5aRs. In future, these insights may be helpful for advancing concepts for therapeutic interventions in inflammatory diseases